Engineering biomaterials to repair damaged or diseased tissues such as cardiac, bone, liver, corneal and skin is an active branch of research in regenerative medicine. One approach being investigated is using cells combined with biomaterial constructs, or scaffolds, that facilitate cell growth and differentiation to create functional tissues in vitro that can be implanted. Three-dimensional (3D) tissue culture systems, which emulate key physical and molecular features of the extracellular microenvironment, provide tremendous advantages to tissue engineering.
Biomaterials may be naturally-derived, such as protein- and polysaccharide-based biomaterials, or synthetic, for example, polymer-, peptide- and ceramic-based biomaterials. Rigorous exploration of properties such as immunogenicity, biodegradability, biocompatibility, ease of modification and permeability is required for the design and development of these biomaterials for clinical tissue engineering. High porosity and adequate pore size, in particular, are important for cell seeding and diffusion of cells and nutrients.
The ready availability and biocompatibility of natural biomaterials such has collagen, alginate and chitosan have made these natural biomaterials attractive substrates for 3D tissue culture. However, challenges with inconsistent mechanical properties and behavior of seeded cells limit their clinical application.
For bone engineering, porosity and pore size of scaffolds have been shown to be a critical factor. Prior studies have indicated that larger pores (˜200-300 μm) result in larger surface area that may promote ion/gas exchange, protein adsorption, and bone apatite mineralization (Karageorgiou et al. Biomaterials 2005 26:5474-91; Yuan et al. Biomaterials 1999 20:1799-806). It is also thought that a larger pore size may be necessary to vascularize implants and mimic the cortical surface and cancellous interior of natural bone (Karageorgiou et al. Biomaterials 2005 26:5474-91). As such, there is a need for scaffolds with large pores that can be seeded with bone cells and used for bone regeneration.
Glucomannan is a naturally-derived polysaccharide composed of a 1:1.6 ratio of β-1,4 linked D-glucose to D-mannose with branches approximately every 11 residues (Alonso-Sande et al. Eur J Phar Biopharm. 2009 72:453-62). Glucomannan has a backbone of approximately 5-10% substituted acetyl groups that participate in hydrogen bonding and hydrophobic interactions that confers solubility. Hydrolysis of the acetyl group in the presence of alkali decreases the solubility of glucomannan and results in aggregation followed by gel formation. Glucomannan is commonly used in foods as an emulsifier or thickener and is being investigated for biopharmaceutical applications due to its gelling and biodegradable properties as well as its malleability to be shaped into films, beads and hydrogels. Glucomannan-based beads, microparticles, and nanoparticles have been developed for DNA and drug delivery (Liu et al. Drug Deliv. 2007 14:397-402; Wang et al. Int J Pharm. 2002 244:117-26; Wen et al. Int J Biol Macromol. 2008 42:256-63) with no significant signs of oral toxicity, skin sensitization, intestinal toxicity, embryotoxicity, or cell-aging observed (Konishi et al. Jpn J Exp Med. 1984 54:139-42).
Glucomannan has recently been investigated as composite scaffolds for chondrocyte culture and injectable scaffolds for cartilage regeneration (Kondo et al. J Tissue Eng Regen Med. 2009 3:361-7). This investigation resulted in the production of a konjac glucomannan/hyaluronic acid hydrogel, wherein cells are cultured and allowed to clump as a suspension in the gel. However, an exploration of developing glucomannan as a porous scaffold for tissue engineering applications has yet to be conducted.
Surprisingly, the present invention provides a glucomannan microporous matrix capable of promoting cell growth and useful as a novel biomaterial scaffold for 3D cell culture and tissue engineering as well as in vivo tissue regeneration, for example, bone regeneration.